The 2',3'-dideoxyriboside of 2,6-diaminopurine selectively inhibits human immunodeficiency virus (HIV) replication in vitro

The 2',3'-dideoxyriboside of 2,6-diaminopurine(ddDAPR) is, like 2',3'-dideoxyadenosine (ddAdo), a potent and selective inhibitor of human immunodeficiency virus (HIV) in vitro. The ddDAPR compound inhibits HIV antigen expression and HIV-induced cytopathogenicity in MT4 cells at a 50% effective dose (ED50) of 2.5-3.6 microM, as compared to 3.1-6.4 microM for ddAdo. Both compounds are endowed with a high selectivity index: 112 for ddDAPR and 139 for ddAdo. The 2',3'-unsaturated derivatives of ddDAPR and ddAdo, i.e. ddeDAPR and ddeAdo, are considerably more cytotoxic and less effective against HIV than the parental compounds. Like ddAdo, ddDAPR is only weakly inhibitory to the proliferation and DNA and RNA synthesis of a series of human B-lymphoblast, T-lymphoblast and T-lymphocyte cell lines. In contrast to ddAdo, which is rapidly deaminated by beef intestine adenosine deaminase at an initial velocity (Vi) of 145 mumol/mg protein/min, ddDAPR and ddeDAPR are poor substrates for the enzyme (Vi: 8 and 0.7 mumol/mg protein/min, respectively), which further contributes to the potential of ddDAPR as a chemotherapeutic agent against AIDS.     

Potent and selective anti-HTLV-III/LAV activity of 2',3'-dideoxycytidinene, the 2',3'-unsaturated derivative of 2',3'-dideoxycytidine

2',3'-Dideoxycytidinene (ddeCyd), the 2',3'-unsaturated derivative of 2',3'-dideoxycytidine (ddCyd) is, like ddCyd itself, a potent and selective inhibitor of HTLV-III/LAV in vitro. This conclusion is based on the relatively high ratio of effective antiviral dose (0.3 microM) versus cell growth inhibitory concentration (20-35 microM) and the lack of any appreciable inhibitory activity against a series of non-oncogenic RNA and DNA viruses. Both compounds were considerably more inhibitory to human lymphoid cell lines than human nonlymphoid or murine cell lines. They were highly dependent on prior activation by deoxycytidine kinase to exert their anti-HTLV-III/LAV and cytostatic effects. In contrast with ddCyd, ddeCyd lost part of its anti-retrovirus effect upon prolonged incubation (10 days) with the virus-infected cells in culture.     

Establishment of mutant murine mammary carcinoma FM3A cell strains transformed with the herpes simplex virus type 2 thymidine kinase gene

To establish cell systems appropriate for investigating the mode of action of antiherpetic nucleoside analogues, mutant cell strains were constructed from murine mammary carcinoma FM3A cells, which were deficient in TK, but were transformed with a recombinant plasmid DNA containing the HSV-2 TK gene. The transformed cells incorporated the viral DNA, expressed viral TK activity and showed unusually high sensitivity to the cytostatic action of the antiherpetic nucleoside analogues ACV and IVDU, both of which were only weakly inhibitory to the growth of the parent cells. Curiously, the FM3A cell strains transformed with HSV-2 TK gene showed a higher sensitivity to ACV and IVDU than the previously established cell line transformed with HSV-1 TK gene. This contrasts with the inhibitory effects of ACV and IVDU on acute HSV infection, since HSV-2 infection is slightly or considerably less susceptible than HSV-1 infection to inhibition by ACV or IVDU, respectively.     

Antiviral activity of the 3''-amino derivative of (E)-5-(2-bromovinyl)-2''-deoxyuridine.

3'-NH2-BV-dUrd, the 3'-amino derivative of (E)-5-(2-bromovinyl)-2'-deoxyuridine, was found to be a potent and selective inhibitor of herpes simplex virus type 1 (HSV-1) and varicella-zoster virus (VZV) replication. 3'-NH2-BV-dUrd was about 4-12 times less potent but equally selective in its anti-herpes activity as BV-dUrd. Akin to BV-dUrd, 3'-NH2-BV-dUrd was much less inhibitory to herpes simplex virus type 2 than type 1. It was totally inactive against a thymidine kinase-deficient mutant of HSV-1. The 5'-triphosphate of 3'-NH2-BV-dUrd (3'-NH2-BV-dUTP) was evaluated for its inhibitory effects on purified herpes viral and cellular DNA polymerases. Among the DNA polymerases tested, HSV-1 DNA polymerase and DNA polymerase alpha were the most sensitive to inhibition by 3'-NH2-BV-dUTP (Ki values 0.13 and 0.10 microM, respectively). The Km/Ki ratio for DNA polymerase alpha was 47, as compared with 4.6 for HSV-1 DNA polymerase. Thus, the selectivity of 3'-NH2-BV-dUrd as an anti-herpes agent cannot be ascribed to a discriminative effect of its 5'-triphosphate at the DNA polymerase level. This selectivity most probably resides at the thymidine kinase level. 3'-NH2-BV-dUrd would be phosphorylated preferentially by the HSV-1-induced thymidine kinase (Ki 1.9 microM, as compared with greater than 200 microM for the cellular thymidine kinase), and this preferential phosphorylation would confine the further action of the compound to the virus-infected cell.     

Molecular cloning of the lectin and a lectin-related protein from common Solomon's seal (Polygonatum multiflorum)

The most prominent protein ofPolygonatum multiflorum (common Solomon's seal) rhizomes has been identified as a mannose-binding lectin. Analysis of the purified lectin demonstrated that it is a tetramer of four identical subunits of 14 kDa. Molecular cloning further revealed that the lectin from this typical Liliaceae species belongs to the superfamily of monocot mannose-binding proteins. Screening of cDNA libraries constructed with RNA isolated from buds, leaves and flowers ofP. multiflorum also yielded cDNA clones encoding a protein, which contains two tandemly arranged domains with an obvious sequence homology to the mannose-binding lectins. Molecular modelling of thePolygonatum lectin and lectin-related protein indicated that the three-dimensional structure of both proteins strongly resembles that of the snowdrop lectin. In addition, this approach suggested that the presumed carbohydrate-binding sites of the lectin can accommodate a mannose residue whereas most of the carbohydratebinding sites of the lectin-related protein cannot.Abbreviations GNA Galanthus nivalis agglutinin - HCA hydrophobic cluster analysis - LECPMA cDNA clone encoding PMA - PM30 30 kDa protein isolated fromPolygonatum multiforum - PMA Polygonatum multiflorum agglutinin - PMLRP Polygonatum multiflorum lectin-related protein     

Intralymphatic administration of BCG in melanoma patients

Summary From November 1973 to December 1974, 20 patients with advanced malignant melanoma were treated with BCG given by intralymphatic route at the Cancer Institute of Milan. The lyophilized Pasteur BCG was used. Patients were treated with a single dose ranging from 0.2–80 mg. Patients' performance status was never severely impaired.The most frequent side effects were fever, lymphangitis, and lymph node enlargement.Variations were observed in white cell count, ERS and immunoglobulins; in no case did we find evidence of liver toxicity or tumor growth enhancement. It is concluded that the intralymphatic route is a safe way of administrating BCG.     

1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (ribavirin) and 5-ethynyl-1-beta-D-ribofuranosylimidazole-4-carboxamide (EICAR) markedly potentiate the inhibitory effect of 2',3'-dideoxyinosine on human immunodeficiency virus in peripheral blood lymphocytes.

Ribavirin and EICAR are two antiviral agents that share a similar antiviral activity spectrum and are targeted at inosine 5'-monophosphate (IMP) dehydrogenase. Neither ribavirin nor EICAR inhibit the replication of human immunodeficiency virus (HIV) in peripheral blood lymphocyte cells (PBL) at subtoxic concentrations. However, both compounds markedly potentiate the anti-HIV activity of 2',3'-dideoxyinosine (DDI) in PBL cells without a marked increase of toxicity. Both the increased IMP levels and the decreased guanine nucleotide levels caused by ribavirin and EICAR may be responsible for their potentiating effect on the anti-HIV activity of DDI.     

New antivirals - mechanism of action and resistance development

In recent years, several novel treatment modalities emerged for a number of virus infections, including lamivudine for hepatitis B virus, abacavir, adefovir dipivoxyl and apropovir disprometil for human immunodeficiency virus, cidofovir for cytomegalovirus, and famciclovir (the oral prodrug of penciclovir) and cidofovir for other herpesviruses (i.e. herpes simplex virus and varicella-zoster virus). For all drugs, resistance eventually develops upon prolonged administration to the infected individuals, albeit at a varying extent. In addition, new mutations related to multidrug resistance have recently been identified.     

6-Azathymidine-4′-thionucleosides: synthesis and antiviral evaluation

The synthesis of dideoxy-6-azathymidine 4′-thionucleoside 1-(2,3-dideoxy-4-thio-β-D-erythro-pentofuranosyl)-(6-azathymidine) (2), and the L-nucleoside, 1-(4-thio-β-L-erythro-pentofuranosyl)-(6-azathymidine) (3) and their evaluation against a wide panel of antiviral assays are described. The L-thionucleoside (3) was devoid of antiviral activity. The dideoxy-thionucleoside (2) was moderately active against vaccinia virus (VV) and the herpes simplex virus strains HSV-1 (strain KOS) and HSV-2 (strain G) (MIC 12 μM) and retained inhibitory activity vs a thymidine kinase-deficient strain HSV-1/TK^–, suggesting that (2) is not dependent on viral TK-catalysed phosphorylation for antiviral activity and/or may use an alternative metabolic activation pathway.     

In vivo Antiretroviral Efficacy of Oral bis(POM)-PMEA,the bis(Pivaloyloxymethyl)prodrug of 9-(2-Phosphonylmethoxyethyl) adenine (PMEA)

Abstract

The bis-pivaloyloxymethyl(POM)- and diphenyl-ester prodrugs of the broad spectrum antiviral agent 9-(2-phosphonylmethoxyethyl)adenine (PMEA) have been evaluated in vivo for antiviral efficacy upon oral administration in severe combined immune deficiency (SCID) mice infected with Moloney murine sarcoma virus (MSV). Oral bis (POM)-PMEA proved highly efficient in delaying MSV-induced tumor formation and associated death, its effect being equal to that of subcutaneous PMEA at an equimolar dose. Compared to bis(POM)-PMEA, oral diphenyl-PMEA had lower antiviral efficacy, whereas PMEA as such was poorly effective when administered orally. Our studies indicate that bis(POM)-PMEA must have a favorable oral bioavailability and justify its clinical investigation as an oral prodrug of PMEA in the treatment of HIV infections.